The goal of this project is to chemically synthesize specific and potent inhibitors of tyrosine-directed protein kinases in vitro. We have chosen the transformation protein of Rous sarcoma virus, pp60src, as our model tyrosine-directed protein kinase. The design of the inhibitor will be peptide or peptide-like and is based on the primary sequence of the active site of pp60src. Our long-range goal will be to synthesize specific, in vivo active inhibitors of tyrosine-directed protein kinase. Such inhibitors are expected to aid studies on the biochemical consequences of the tyrosine-specific phosphorylation as a cellular regulation in tumor transformation and growth. It will also provide a prototype drug for the in vivo regulation of tumor growth. To aid our studies on the design of pp60srcprotein kinase inhibitors and to further our understanding of the mechanism of pp60srcas a protein kinase, our short-range goals also include: (1)\the purification of pp60src; (2) the study of the functional domains of pp60src, particularly the probable ATP-binding site and the phosphotyrosine active site; (3) the investigation of the phosphotransferring property and chemical reactivity of the high-energy O-phosphate-tyrosine; and (4) enzyme kinetic studies of substrates and inhibitors of pp60src. (B)